U.S. patent application number 15/469745 was filed with the patent office on 2017-11-23 for processes of treating grain.
The applicant listed for this patent is Dupont Industrial Bioscience, The United States of America, as Represented by the Secretary of Agriculture. Invention is credited to David Johnston, Bart C. Koops, Jayarama K. Shetty.
Application Number | 20170332663 15/469745 |
Document ID | / |
Family ID | 60001393 |
Filed Date | 2017-11-23 |
United States Patent
Application |
20170332663 |
Kind Code |
A1 |
Johnston; David ; et
al. |
November 23, 2017 |
Processes of Treating Grain
Abstract
Processes of treating grain (e.g., corn), involving milling the
grain to produce milled grain wherein the grain germ remains intact
in the milled grain, and producing a mixture by mixing the milled
grain with water and at least one enzyme selected from the group
consisting of protease, alpha amylase, glucoamylase, cell wall
degrading enzyme, and mixtures thereof, wherein the pH of the
mixture is optionally adjusted to a pH of about 3.5 to about 6.5,
and incubating the mixture for about 1 to about 3 hours to produce
an incubated mixture.
Inventors: |
Johnston; David; (Wyndmoor,
PA) ; Shetty; Jayarama K.; (Pleasanton, CA) ;
Koops; Bart C.; (Zuid Holland, NL) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
The United States of America, as Represented by the Secretary of
Agriculture
Dupont Industrial Bioscience |
Washington
Palo Alto |
DC
CA |
US
US |
|
|
Family ID: |
60001393 |
Appl. No.: |
15/469745 |
Filed: |
March 27, 2017 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
62317807 |
Apr 4, 2016 |
|
|
|
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
B02B 3/00 20130101; A23K
10/12 20160501; A23K 10/30 20160501; Y02E 50/16 20130101; Y02E
50/17 20130101; C12P 7/10 20130101; C12P 2201/00 20130101; C12Y
304/00 20130101; A23K 10/14 20160501; Y02E 50/10 20130101; B02C
11/08 20130101; C12P 19/02 20130101; C12P 19/14 20130101 |
International
Class: |
A23K 10/12 20060101
A23K010/12; B02B 3/00 20060101 B02B003/00; A23K 10/30 20060101
A23K010/30; B02C 11/08 20060101 B02C011/08; A23K 10/14 20060101
A23K010/14 |
Claims
1. A process of treating grain, said process comprising milling
said grain to produce milled grain wherein the grain germ remains
intact in said milled grain, and producing a mixture by mixing said
milled grain with water and at least one enzyme selected from the
group consisting of protease, alpha amylase, glucoamylase, cell
wall degrading enzyme, and mixtures thereof, wherein the pH of said
mixture is optionally adjusted to a pH of about 3.5 to about 6.5,
and incubating said mixture for about 1 to about 3 hours to produce
an incubated mixture.
2. The process of claim 1, further comprising separating said
incubated mixture into germ and a liquid slurry, reducing the
particle size of materials in said liquid slurry by grinding,
adding yeast and amylase to said liquid slurry and fermenting for
about 30 to about 72 hours, removing ethanol from said liquid
slurry and separating said liquid slurry into fiber and a defibered
liquid slurry, and separating said defibered liquid slurry to
produce protein solids.
3. The process of claim 1, further comprising separating said
incubated mixture into germ and a liquid slurry, reducing the
particle size of materials in said liquid slurry by grinding,
adding amylase to said liquid slurry to hydrolyze the starches in
said liquid slurry, and separating said liquid slurry into fiber
and a defibered liquid slurry, and separating said defibered liquid
slurry into protein solids and a solution containing sugars.
4. The process of claim 1, further comprising separating said
incubated mixture into germ and a liquid slurry, reducing the
particle size of materials in said liquid slurry by grinding,
adding amylase to said liquid slurry and heating for about 15 min
to about 2 hours, adding glucoamylase enzyme to said liquid slurry
to hydrolyze the starches in said liquid slurry, and separating
said liquid slurry into fiber and a defibered liquid slurry, and
separating said defibered liquid slurry into protein solids and a
solution containing sugars.
5. The process of claim 1, wherein said grain is corn.
Description
REFERENCE TO RELATED APPLICATION
[0001] This application claims the benefit of U.S. Provisional
Application No. 62/317,807, filed 4 Apr. 2016, which is
incorporated herein by reference in its entirety.
BACKGROUND OF THE INVENTION
[0002] Disclosed are processes of treating grain (e.g., corn),
involving milling the grain to produce milled grain wherein the
grain germ remains intact in the milled grain, and producing a
mixture by mixing the milled grain with water and at least one
enzyme selected from protease, alpha amylase, glucoamylase, cell
wall degrading enzyme, or mixtures thereof, wherein the pH of the
mixture is optionally adjusted to a pH of about 3.5 to about 6.5,
and incubating the mixture for about 1 to about 3 hours to produce
an incubated mixture.
[0003] The majority of fuel ethanol plants today currently utilize
similar processes to produce Distillers Dried Grains with Solubles
(DDGS), some produce liquid carbon dioxide, and most produce
post-fermentation corn oil as the only coproducts. Several plants
have modified processes to utilize a dry milling fractionation
process prior to fermentation for recovery of germ and pericarp
fiber. A few plants have wet separation processes that allow the
production of a protein rich feed (about 50% protein) with
relatively low fiber content as a new coproduct. Most of the other
variations are strictly towards process improvements and do not
produce additional new coproducts.
[0004] Removal of germ and pericarp fiber using a dry milling
fractionation process prior to ethanol fermentation has energy
saving advantages over the conventional process because the
components removed are not wetted before separation and removal.
However, the dry fractionation process unfortunately removes some
essential nutrients needed for efficient fermentation with the germ
fraction and can lose a significant amount of starch if processing
is not done properly. The germ is also relatively low in oil
concentration (.apprxeq.18%) because it retains these soluble
components; additionally, the pericarp fiber is not by itself a
particularly valuable co-product.
[0005] Wet fractionation processes such as quick germ/fiber or
E-Mill processing require relatively more energy for drying but do
not lose fermentation nutrients or as much starch relative to dry
fractionation (Singh, V., and S. R. Eckhoff, Cereal Chemistry,
73(6): 716-720 (1996); Singh, V., et al., Cereal Chemistry, 82(2):
187-190 (2005); Murthy, G. S., et al., Cereal Chemistry, 83(5):
455-459 (2006); Murthy, G. S., et al., Industrial Biotechnology,
7(4): 298-307 (2011); U.S. Pat. No. 6,899,910). The germ is higher
in oil concentration relative to dry milled germ, making it as
valuable as germ from the wet milling process. In both germ
recovery processes it is essential that the germ is not broken
during the milling processes or oil can be lost.
[0006] In the dry process, tempering of the kernel (which raises
the moisture content of the kernel) prior to grinding in a special
mill is required. There are also sizing and air classification
steps taken to separate components after grinding. With the
existing wet fractionation processes, the whole kernels need to be
soaked in water from about 3 to about 12 hours prior to grinding in
a toothed disk mill. This keeps the germ from breaking during
milling and helps remove endosperm (and starch) from the separated
fractions. The hydration also results in the leaching of the
soluble components from the germ that aid in fermentation and help
concentrate the oil within the germ. The loss of these soluble
components during the dry fraction process results in sluggish or
stuck fermentations, and has forced plants utilizing dry
fractionation to incompletely recover germ and/or to add
supplemental nutrients in order to alleviate these problems.
[0007] Several processes that combine dry fractionation with wet
milling have also been developed (U.S. Pat. No. 3,597,274; U.S.
Pat. No. 4,181,748). The goals of these processes were to isolate
starch by first dry milling the corn and then treating all or part
of the ground kernel with sulfur dioxide in order to isolate
starch. These processes resulted in significant starch losses that
would be unacceptable from an economic standpoint if applied in an
ethanol process.
[0008] Work done in our laboratory on the development of the
E-Milling (Enzymatic Wet Milling) process (U.S. Pat. No. 6,566,125;
Johnston, D. B., and V. Singh, Cereal Chemistry, 78(4): 405-411
(2001)) showed that the addition of a protease could replace the
sulfite chemical processing and release starch for separation. We
later expanded on this technology for the development of the germ
and pericarp fiber fractionation process (E-Mill) for use with
ethanol production that could utilize other enzymes in addition to
proteases (Johnston, D. B., et al., JAOCS, 82(8): 603-608 (2005);
Singh et al, 2005). With both of these processes, it was necessary
to break the corn kernel without disrupting the germ before the
enzyme treatment would be effective. This mandated some form of
soaking process be utilized prior to grinding with a toothed disc
mill. The capital cost of implementing the soaking step is one of
the reasons the wet fraction processes have not found broad
acceptance. Reduction of the initial capital costs or significant
reduction in operating costs could result in broad adoption of wet
fractionation technology in the fuel ethanol industry.
[0009] We have developed a simplified enzymatic germ and fiber
(pericarp and endosperm) recovery process for use in an ethanol
production system that could be implemented using less capital
relative to the E-Mill or Quick Germ/Fiber processes and
surprisingly does not have the significant ethanol loss issues of
other processes.
SUMMARY OF THE INVENTION
[0010] Disclosed are processes of treating grain (e.g., corn)
involving milling the grain to produce milled grain wherein the
grain germ remains intact in the milled grain, and producing a
mixture by mixing the milled grain with water and at least one
enzyme selected from the group consisting of protease, alpha
amylase, glucoamylase, cell wall degrading enzyme, and mixtures
thereof, wherein the pH of the mixture is optionally adjusted to a
pH of about 3.5 to about 6.5 (e.g., 3.5 to 6.5), and incubating the
mixture for about 1 to about 3 hours (e.g., 1 to 3 hours) to
produce an incubated mixture.
[0011] The processes may further involve separating the incubated
mixture into germ and a liquid slurry, reducing the particle size
of materials in the liquid slurry by grinding, adding yeast and
amylase to the liquid slurry and fermenting for about 30 to about
72 hours (e.g., 30 to 72 hours), removing ethanol from the liquid
slurry (e.g., by distillation) and separating (e.g., screening) the
liquid slurry into fiber and a defibered liquid slurry, and
separating (e.g., centrifuging) the defibered liquid slurry to
produce protein solids.
[0012] The process may further involve separating the incubated
mixture into germ and a liquid slurry, reducing the particle size
of materials in the liquid slurry by grinding, adding amylase to
the liquid slurry to hydrolyze the starches in the liquid slurry,
and separating (e.g., screening) the liquid slurry into fiber and a
defibered liquid slurry, and separating (e.g., centrifuging) the
defibered liquid slurry into protein solids and a solution
containing sugars.
[0013] The process may further involve separating the incubated
mixture into germ and a liquid slurry, reducing the particle size
of materials in the liquid slurry by grinding, adding amylase to
the liquid slurry and heating (e.g. about 60.degree. to about
130.degree. C. (60-130.degree. C.), preferably about 75.degree. to
about 105.degree. C. (75-105.degree. C.)) for about 15 min to about
2 hours (e.g., 15 min to 2 hours) (liquefaction), adding
glucoamylase enzyme to the liquid slurry to hydrolyze the starches
in the liquid slurry (saccharification), and separating (e.g.,
screening) the liquid slurry into fiber and a defibered liquid
slurry, and separating (e.g., centrifuging) the defibered liquid
slurry into protein solids and a solution containing sugars.
[0014] This summary is provided to introduce a selection of
concepts in a simplified form that are further described below in
the detailed description. This summary is not intended to identify
key features or essential features of the claimed subject matter,
nor is it intended as an aid in determining the scope of the
claimed subject matter.
BRIEF DESCRIPTION OF THE DRAWINGS
[0015] FIG. 1 shows particle size analysis of corn ground using a
degermination mill for untempered (shaded), ground after tempering
and sized wet (dotted), and ground after tempering and dried
(diagonal stripes) prior to sizing as described below. Bars are
averages of duplicate milling runs and data labels represent the
percent of total oil content for each fraction.
[0016] FIG. 2A and FIG. 2B are flow charts showing major steps for
the degermination process and the conventional process as described
below. Inputs are shown on the left and outputs on the right. FIG.
2C and FIG. 2D are flow charts showing major steps for the
degermination process for the production of a sugar solution as
described below. FIG. 2C shows the process without thermal
gelatinization of the starch, and FIG. 2D shows the process with
the thermal treatment and the separate additions of alpha amylase
and glucoamylase.
[0017] FIG. 3 shows average theoretical weight loss for
fermentation done with germ recovery (dotted line) and for
conventional fermentations (solid line) as described below.
[0018] FIG. 4 shows fraction yields for components relative to
starting corn as described below. The DDGS equivalents (dotted),
Germ (dark shaded) and protein fractions (light shaded) are shown
as the averaged values made on a dry weight basis. Error bars
represent .+-.one standard deviation of the average.
[0019] FIG. 5A shows oil yields for fraction relative to the total
weight recovered and FIG. 5B shows protein yields for fraction
relative to the total weight recovered as described below. The DDGS
equivalents (dotted), Germ (dark shaded), and protein fractions
(light shaded) are shown as the averaged values made on a dry
weight basis. Error bars represent .+-.one standard deviation of
the average.
DETAILED DESCRIPTION OF THE INVENTION
[0020] Disclosed are processes of treating grain (e.g., corn),
involving milling the grain to produce milled grain wherein the
grain germ remains intact in the milled grain, and producing a
mixture by mixing the milled grain with water and at least one
enzyme selected from the group consisting of protease, alpha
amylase, glucoamylase, cell wall degrading enzyme, and mixtures
thereof, wherein the pH of the mixture is optionally adjusted to a
pH of about 3.5 to about 6.5 (e.g., 3.5 to 6.5), and incubating the
mixture for about 1 to about 3 hours (e.g., 1 to 3 hours) to
produce an incubated mixture.
[0021] The processes may further involve separating the incubated
mixture into germ and a liquid slurry, reducing the particle size
of materials in the liquid slurry by grinding, adding yeast and
amylase to the liquid slurry and fermenting for about 30 to about
72 hours (e.g., 30 to 72 hours), removing ethanol from the liquid
slurry (e.g., by distillation) and separating (e.g., screening) the
liquid slurry into fiber and a defibered liquid slurry, and
separating (e.g., centrifuging) the defibered liquid slurry to
produce protein solids.
[0022] The process may further involve separating the incubated
mixture into germ and a liquid slurry, reducing the particle size
of materials in the liquid slurry by grinding, adding amylase to
the liquid slurry to hydrolyze the starches in the liquid slurry,
and separating (e.g., screening) the liquid slurry into fiber and a
defibered liquid slurry, and separating (e.g., centrifuging) the
defibered liquid slurry into protein solids and a solution
containing sugars.
[0023] The process may further involve separating the incubated
mixture into germ and a liquid slurry, reducing the particle size
of materials in the liquid slurry by grinding, adding amylase to
the liquid slurry and heating (e.g., about 60 to about 130.degree.
C. (60-130.degree. C.), preferably about 75 to about 105.degree. C.
(75-105.degree. C.)) for about 15 min to about 2 hours (e.g., 15
min to 2 hours) (liquefaction), adding glucoamylase enzyme to the
liquid slurry to hydrolyze the starches in the liquid slurry
(saccharification), and separating (e.g., screening) the liquid
slurry into fiber and a defibered liquid slurry, and separating
(e.g., centrifuging) the defibered liquid slurry into protein
solids and a solution containing sugars.
[0024] Utilizing a dry grinding system, corn (or other grains) can
be ground so that the germ is not broken or damaged. This can be
done using tempered or untempered grain; tempering generally is
conducted at about 60.degree. C. or below for about 15 minutes to
about 3 hours. The grain is then mixed in water at a temperature
below the gelatinization temperature of the starch (the
gelatinization temperature of corn starch is typically about
70.degree. C. but it can be higher or lower depending on the
specific type of corn used; generally the range could be as low as
62.degree. C. and as high as 78.degree. C.), and the pH adjusted to
the active range of the specific enzyme(s) being used. The grain is
then incubated for a period of about 1 to about 6 hours (e.g., 1 to
6 hours) so that the specific gravity of the suspension is
sufficient for the germ to float and be recovered. If the specific
gravity is sufficiently increased, the coarse fiber could also
float and be recovered with the germ. This recovery could be done
using hydrocyclones or by floatation. Once the germ (and
potentially the coarse fiber) has been removed, the particle sizes
of the remaining solids in the slurry are reduced in size using an
appropriate grinding device. The slurry can then be further
incubated and or treated with other enzymes and used to produce
several different products.
[0025] The general ethanol production process is well known in the
art. Ethanol can be made by incubating with either native starch
degrading enzymes or with thermostable starch degrading enzymes
along with an elevated temperature treatment above the
gelatinization temperature of the starch. The solution is then
fermented with yeast to produce ethanol. After fermentation, the
ethanol is removed by distillation. Following distillation, the
fine fiber is recovered using fine mesh screens. The insoluble
protein and yeast cells can then be recovered separately from the
liquid stream by centrifugation or fine filtration such as vacuum
belt filtration or micro filtration membranes. Non-germ oil can
optionally be recovered using a high-speed centrifugation.
[0026] Starch can be isolated if the enzyme used is a protease that
can break down the protein matrix surrounding the starch granules,
and the finely ground slurry is further incubated to release the
starch granules (Enzymatic Wet Milling process). Following this
treatment, the slurry can then be separated using standard wet
milling separation systems for starch, gluten and fiber
recovery.
[0027] A sugar solution may also be prepared by treating with a
combination of native starch degrading enzymes under temperature
and enzyme combinations that can hydrolyze the starch and release
glucose, maltose or a combination of oligosaccharides from starch
hydrolysis. Following this hydrolysis, the fiber and insoluble
protein can be separated as above and the fiber and insoluble
protein washed to remove residual sugars. The sugar solution can
then be utilized for ethanol production or other purposes. The
solution can also be treated to remove the insoluble protein by
centrifugation or fine filtration such as vacuum belt filtration or
micro filtration membranes.
[0028] Utilizing this system, corn or other grains (e.g., sorghum,
wheat, barley, rye and triticale) can be enzymatically processed
without the need for soaking the grain prior to grinding. This will
save significantly on capital cost for implementation and reduce
processing times.
[0029] "Optional" or "optionally" means that the subsequently
described event or circumstance may or may not occur, and that the
description includes instances in which said event or circumstance
occurs and instances where it does not. For example, the phrase
"optionally comprising a defoaming agent" means that the
composition may or may not contain a defoaming agent and that this
description includes compositions that contain and do not contain a
foaming agent.
[0030] By the term "effective amount" of a compound or property as
provided herein is meant such amount as is capable of performing
the function of the compound or property for which an effective
amount is expressed. As will be pointed out below, the exact amount
required will vary from process to process, depending on recognized
variables such as the compounds employed and the processing
conditions observed. Thus, it is not possible to specify an exact
"effective amount." However, an appropriate effective amount may be
determined by one of ordinary skill in the art using only routine
experimentation.
[0031] Unless defined otherwise, all technical and scientific terms
used herein have the same meaning as commonly understood by one of
ordinary skill in the art to which the invention belongs. As used
herein, the term "about" refers to a quantity, level, value or
amount that varies by as much as 10% to a reference quantity,
level, value or amount. Although any methods and materials similar
or equivalent to those described herein can be used in the practice
or testing of the present invention, the preferred methods and
materials are now described.
[0032] The following examples are intended only to further
illustrate the invention and are not intended to limit the scope of
the invention as defined by the claims.
Examples
[0033] Materials and Enzymes:
[0034] The enzymes used were gifts of DuPont Industrial Biosciences
(Palo Alto, Calif.). SPEZYME RSL.RTM. (thermostable
.alpha.-amylase), OPTIDEX.RTM. L-400 (glucoamylase), PROSTEEP.RTM.
(a protease), and FERMGEN.RTM. (an acid protease) were all
commercially available products and used by volume with the
activities as defined by the manufacturer. The Red Star Ethanol Red
yeast was a gift from Lesaffre (Milwaukee, Wis.). Urea was from IBI
Scientific (Peosta, Iowa). Corn was #2 yellow dent commodity corn
grown during the 2014 crop year.
[0035] Corn Grinding and Analysis:
[0036] Corn was ground for germ recovery experiments without
tempering using a custom-built degermination mill; however,
commercially available mills (off the shelf) could readily be used.
The mill utilized a rotary impact grinding mechanism surrounded by
a screen with 9.65 mm (0.38'') diameter round holes. The mill is
described in detail by Brekke, O. L., et al. (Cereal Chemistry
48(5): 499-511 (1971)). Our mill was built at the USDA Eastern
Regional Research Center, Wyndmoor, Pa.
[0037] For experiments without germ recovery, corn was ground using
a Bunn (Springfield, Ill.) model G2 burr mill coffee grinder. The
gap was set so that the ground corn produced would pass through a 1
mm screen.
[0038] The fermentations were done using 553 grams of corn that was
weighed prior to grinding and the entire amount collected directly
into the tared beakers. A smaller quantity of the same corn was
ground and collected for moisture determinations. Moisture content
was determined each day and for each grinding method used according
to AOAC Official Method 930.15 (AOAC International, 2005). The
measured dry weights were used for all mass balance and yield
determinations.
[0039] Conventional Mash Preparation with Additional Particle Size
Reduction:
[0040] The ground corn (still containing the corn germ but is
ground intact) was mixed with water in a tared 2 L beaker and the
total slurry weight adjusted to 1500 grams. The particle size of
the slurry was further reduced using an IKA (Wilmington, N.C.) T25
Disperser with an 18G dispersing element at 16,000 rpm for 3 to 5
min until the slurry was uniform. A mechanical mixer was used and
the pH adjusted to 5.8 using 1 N HCl. Alpha amylase (SPEZYME
RSL.RTM.) was added at 0.5 mL/kg mash and the slurry heated on a
hot plate and held for 60 min at 95.degree. C. The mechanical mixer
speed was adjusted as necessary as the slurry viscosity changed to
maintain a well-mixed slurry. The mash was cooled in a water bath
to 30.degree. C. and supplemental N (400 ppm) added as urea. The pH
was adjusted to 4.5 with 1 N HCl and glucoamylase (OPTIDEX.RTM.
L-400) was added at a dosage of 0.4 mL/kg of mash. Active yeast was
added (Red Star Ethanol Red; 1.1 g/kg of mash) to start the
fermentation and the entire slurry transferred into a tared 2 L
Erlenmeyer flask. An acid protease (FERMGEN.RTM.) was added at 0.67
ml/kg of mash and the total slurry weight was then readjusted to
1500 g with water to compensate for evaporation losses. Small
amounts of water for rinsing were required throughout the
processing for probe and mixer rinsing and for final transfer to
the incubation flask; however, these were less than what was lost
through evaporation while heating. The flask was stoppered and a
21-gauge needle inserted to vent CO.sub.2 produced.
[0041] Germ Recovery Mash Preparation:
[0042] The degerminated ground corn (with intact corn germ) was
mixed with water in a 2 L beaker and the pH adjusted to 4.5 using 1
N HCl. PROSTEEP.RTM. was added at 0.67 g/kg of mash and the total
slurry weight adjusted to 1500 grams. The beaker was then incubated
in a water bath at 48.degree. C. and mixed with a mechanical mixer
for 2 hours. Following incubation, the beaker was removed and the
floating germ recovered by skimming using a small section of 2 mm
wire screen (#10 mesh). The slurry was periodically mixed to allow
more germ to float and again skimmed. This was done repeatedly
until no more germ could be recovered (about 20 min). The germ was
rinsed with 100 mL of water and the rinse water added to the
degermed slurry. The germ collected was dried at 55.degree. C. in a
forced air oven. The degermed corn slurry was readjusted to 1500 g
with water and processed without adjusting the pH (now about pH
4.9). The particle size of the slurry was further reduced using an
IKA (Wilmington, N.C.) T25 Disperser with an 18G dispersing element
at 16,000 rpm for 3-5 min until the slurry was uniform. A
mechanical mixer was used and Alpha amylase (SPEZYME RSL.RTM.) was
added at 0.5 mL/kg mash and the slurry heated on a hot plate and
held for 60 min at 95.degree. C. The mechanical mixer speed was
adjusted as necessary as the slurry viscosity changed to maintain a
well-mixed slurry. The mash was cooled in a water bath to
30.degree. C. and supplemental N (400 ppm) added as urea.
Glucoamylase (OPTIDEX.RTM. L-400) was added at a dosage of 0.4
mL/kg of mash. Active yeast was added (Red Star Ethanol Red; 1.1
g/kg of mash) to start the fermentation and the entire slurry
transferred into a tared 2 L Erlenmeyer flask. The total slurry
weight was then readjusted to 1500 g with water to compensate for
evaporation losses. Small amounts of water for rinsing were
required throughout the processing for probe and mixer rinsing and
for final transfer to the incubation flask; however, these were
less than what was lost through evaporation while heating. The
flask was stoppered and a 21-gauge needle inserted to vent CO.sub.2
produced.
[0043] Fermentation Rate and HPLC Analysis:
[0044] Flasks were incubated in a New Brunswick Scientific
Innova.RTM. 44 incubator at 30.degree. C. with shaking at 200 rpm.
Flasks were periodically removed and weighed to determine the
weight loss due to CO.sub.2 production and for fermentation rate
analysis. Flasks were incubated for up to 72 hours. A small sample
was taken at the end of the fermentation for HPLC determination of
maltodextrins (DP4+), maltotriose (DP3), maltose, glucose,
fructose, succinic acid, lactic acid, acetic acid, glycerol,
methanol, and ethanol (Johnston, D. B. and A. J. McAloon,
Bioresource Technology, 154: 18-25 (2014)). The starch content of
the corn was measured using AACC Method 76-13 (AACC International
2000) and used to determine ethanol yields.
[0045] Fiber, Protein and DDGS Recovery:
[0046] Following fermentation, the entire contents of the flask
were transferred to a 2 L beaker and heated to 85.degree. C. with
mixing to remove the ethanol. Following ethanol removal, the fiber
was recovered by screening through a 325 mesh (45 micron) screen
(Dual manufacturing Co. Chicago, Ill.). The fiber was dewatered on
the screen as much as possible by pressing with a spatula. No
washing of the fiber was done at this stage. The volume of the
screened stillage was measured, a sample taken for total solids
analysis and then transferred into centrifuge bottles. The bottles
were centrifuged in a bench top centrifuge at 2000.times.g for 10
min. The bottles were decanted and the volume of liquid measured
and again a sample was taken for total solids analysis. The pellet
of primarily insoluble protein and yeast cells was recovered from
the centrifuge bottles and transferred with a small amount of DI
water used for rinsing into tared weigh boats and dried at
55.degree. C. The recovered fiber was mixed with the stillage and
the mixture was freeze dried to produce a DDGS equivalent.
Fermentations not requiring protein recovery were freeze dried
after the ethanol removal step.
[0047] A washing procedure was also evaluated where the decanted
liquid was added back to the fiber and rescreened through the 325
mesh screen. The liquid was then centrifuged again and the
recovered pellet added to the protein solids. This was repeated two
additional times followed by a 500 mL water wash.
[0048] Compositional Analysis:
[0049] Moisture analysis was done according to AOAC Official Method
930.15. Protein was determined by combustion analysis according to
AOAC 99.03. Crude lipid was determined using AOAC 945.16 or as
described in Johnston et al. 2005 using a Dionex ASE extractor
(AOAC International, 2012). Fiber analysis (ADF and NDF) was done
using an ANKOM fiber analyzer and the manufacturer standard
protocols. Crude Fiber was done using AOCS approved procedure Ba
6a-05. Starch was analyzed using AACC 76-11 with a YSI analyzer for
glucose determination. All conditions and analytical testing were
done in duplicate and results reported as averages.
[0050] Results and Discussion.
[0051] Process Development and Grinding Characterization:
Preliminary fermentation experiments were conducted using
degermination ground corn and compared with conventional grinding.
The results showed slow fermentations that took up to 5 days to
reach completion. Ethanol yields were found to be significantly
reduced when compared to the conventionally ground corn at 72
hours. These experiments led to the implementation of the secondary
size reduction step using a wet grind process shown in FIG. 2A.
This was also incorporated in the conventional process to eliminate
any variation due to the added particle size reduction.
[0052] In dry milling, tempering of the corn kernels (which raises
the moisture content of the kernel) is done prior to grinding. This
aids in the separation and fractionation of the components. Since
our use of the mill was only to break the kernels without damaging
the germ, we conducted all of our fermentations and recovery
experiments without tempering; however, a comparison was done to
evaluate the oil content of fractions with and without tempering
and for particle size analysis. FIG. 1 shows the particle
distribution for corn ground untempered (as used in germ recovery
experiments), tempered and sieved undried (wet), and tempered and
dried before sieving. Sieves having 3.35, 2.36, 2.00, 0.710 mm
openings and pan (#6, 8, 10, 25, and pan) were chosen to
differentiate larger particle sizes. Tempering was done at
60.degree. C. for 60 minutes. The final moisture content was found
to be 25.75% for the tempered corn and 12.76% for the untempered
corn. The particle distribution clearly shows that the grind was
much coarser than typically used in commercial ethanol facilities
(Rausch, K. D., et al., Transactions of the ASAE, 48(1): 273-277
(2005)). Over 70% of the fractions were 2 mm or larger whereas
typically over 85% would pass through this screen. The tempered
corn had noticeably larger pieces of pericarp fiber relative to the
untempered sample.
[0053] Batches of 100 kernels were ground and hand sorted in order
to visually evaluate germ quality. No intact kernels and primarily
intact germ with little apparent damage were surprisingly found.
Undamaged germ is important for retaining the oil content within
the germ during wet fractionation. Oil is known to be present
outside of the germ in other parts of the kernel; however, the
majority of oil in the kernel is located inside of the germ
(80-85%; Watson, S. A., Description, Development, Structure, and
Composition of the Corn Kernel, IN Corn chemistry and Technology,
P. A. White and L. A. Johnston, 2003, pp. 69-106).
[0054] The sized fractions were analyzed for oil content and the
percent of total kernel oil for each fraction is shown in FIG. 1 as
data labels at the top of the bars. The results indicate that more
than 85% of the total kernel oil was present in the two largest
size fractions (>2.36 mm) that visually contain the intact germ.
The tempered sample, that was fractionated prior to drying, showed
86.1% of the total oil in the >3.35 mm fraction with only 8.7%
in the >2.36 mm fraction. When the tempered corn was dried after
grinding prior to sieving, the oil content of the two largest
fractions surprisingly more closely matched the untempered corn
samples. This surprisingly indicated that the germ was still
primarily intact, when ground with or without tempering using this
type of mill and had the potential to be recovered in the germ
fraction. It should be noted that the untempered fractions did have
a larger percentage of oil in the two smallest fractions relative
to either tempered sample, possibly indicating some germ being
damaged.
[0055] Preliminary experiments were also done to evaluate the
incubation time and the protease enzyme dose required for optimal
germ recovery. Incubation times from 1-6 hours, temperatures from
40-60.degree. C., and enzyme doses from 0.5-6 g/kg of corn were
examined for germ recovery yields. It was surprisingly found that
acceptable germ yields were reached by incubating for 2 hours at
48.degree. C. with 2 g/kg of enzyme. Oil concentrations could be
somewhat improved using longer incubation times; however, this
complicated experimentations due to overall processing time. Lower
pH and higher temperatures allowed reduced enzyme dosing but
required base addition for raising the pH after germ recovery or
the alpha amylase used was not sufficiently active. The future use
of a low pH alpha amylase would be of significant benefit as no
additional pH adjustment would be required.
[0056] It is important to note that many other types of enzymes,
not just protease, could be utilized for the germ recovery process.
In U.S. Pat. No. 6,899,910 (Johnston & Singh (2005)), we
demonstrated that a range of enzymes could be utilized to hydrolyze
endogenous components and alter the specific gravity to allow germ
recovery. These other types of enzymes include amylases, proteases,
and cell wall degrading enzyme or mixtures of these enzymes as well
as other enzymes that could hydrolyze endogenous components to
increase the specific gravity.
[0057] Process Description:
[0058] FIG. 2A shows the germ recovery processing flow used for
experiments and FIG. 2B shows the conventional process used when
germ was not recovered. Each also shows the alternative process
where conventional DDGS or Degermed DDGS are produced without fiber
screening or protein recovery.
[0059] Fermentation Rate Comparison:
[0060] Previous research has shown that fermentation rates and
ethanol yields can be affected by the addition of protease during
fermentation (Johnston and McAloon 2014; U.S. Pat. No. 5,231,017
(1991): Wang et al., Cereal Chem. 84(1): 10-14 (2007)). The process
being described here also incorporated a secondary grinding that
significantly reduced the particle size relative to the normal
process and could surprisingly result in altering the rate and/or
yield. To eliminate this as a variable, the secondary grinding step
was also incorporated into the conventional process shown in FIG.
2B.
[0061] Following the weight loss of the flasks over time was done
to compare the fermentation rates and for overall mass balance.
FIG. 3 shows the average weight loss curves for the conventional
process and for our germ recovery process (dotted curve) as a
percentage of the theoretical. Our germ recovery process
surprisingly initially fermented somewhat faster relative to the
conventional process; this is partially the result of reduced
solids in the fermentation flask due to germ removal. Final weight
loss using the germ recovery step was surprisingly only reduced by
2.3% relative to the conventional process where germ was still
present during fermentation; this was likely due to some starch
being lost with the germ fraction. Additional washing of the germ
could reduce this loss; however, it was not possible with the
laboratory scale batch process without significantly reducing the
final solids content. In a continuous operating ethanol plant, the
process condensate could first be used to wash the germ prior to
being added to the ground corn. Some starch loss with germ recovery
will be unavoidable but surprisingly it could potentially be as low
as 1%.
[0062] Fraction Yields:
[0063] The average germ recovery yield using protease was
surprisingly found to be 8.06% of the starting corn on a dry weight
basis. Preliminary trials using lower concentrations of protease
reduced yields or required longer incubation time. This recovery
value was surprisingly consistent with values for germ recovery
yields from wet milling as well as other wet fractionation
processes (Johnston et al. 2005; Eckhoff, S. R., et al., Cereal
Chemistry, 70(6): 723-727 (1993); Ramirez, E. C., et al.,
"Enzymatic corn wet milling: Engineering process and cost model,"
Biotechnology for Biofuels, 2(2): article no. 2 (2009); Singh et
al. 1996; Singh, V., et al., Cereal Chem., 82: 187-190 (2005))).
Alpha amylase (Spezyme RSL) was also tested for germ recovery in
place of the protease. Germ recovery surprisingly was slightly
reduced to 7.2% in these experiments; however, conditions were not
optimized and yield could likely be improved. Using the alpha
amylase prior to liquefaction would surprisingly remove the
additional cost for protease.
[0064] Surprisingly the time needed for germ recovery was
significantly less than needed for other wet fractionation
processes (generally about 2 hours, shorter times could work by
increasing the amount of enzyme used, but the germ is not as high a
quality, fewer solubles leach out of the germ and as a consequence
the oil content is reduced). Conventional corn wet milling requires
24-36 hours of steeping, quick germ utilizes up to 12 hours of
soaking, and the E-Mill process utilizes up to 6 hours of combined
soaking and incubation before germ can be recovered (Blanchard, P.
H., Technology of Corn Wet Milling and Associated Process, 1992,
Industrial Chemistry Library, Elsevier, Amsterdam, The Netherlands;
Singh et al. 1996, 2005). Conventional dry milling (dry
fractionation) is faster; however, the low oil concentration of the
germ, the high starch loss, and reduced fermentation rates make it
a less desirable process.
[0065] Fiber recovery was done in conjunction with germ recovery,
with protein recovery, and with germ and protein recovery to
compare yields and final composition. The solubles recovered were
added to the fiber fraction to produce DDGS equivalents. The yields
were compared with control values without recovery of germ or
protein and the yields are shown in FIG. 4. Surprisingly germ
removal alone reduced the DDGS equivalent yield 24.5% relative to
the control and protein recovery alone reduced the yield 14.5%. The
combined removal of germ and protein surprisingly reduced the yield
39.4% relative to the control. Washing of the fiber as described
above surprisingly resulted in an additional decrease of 2%
reaching a 41.5% reduction in DDGS equivalents relative to the
control. Surprisingly there was a corresponding increase in the
protein fraction.
[0066] The post-fermentation recovery of fiber surprisingly reduced
the amount of starch lost relative to previous processes that
implemented fiber recovery prior to fermentation; however, it does
not increase fermenter space because the fiber is still present
during fermentation (by removing things prior to fermentation (as
with up front germ recovery) the amount of non-fermentable material
going into the fermenter is reduced thereby increasing fermenter
space). Surprisingly germ recovery prior to fermentation in the
current process did significantly increase fermentation space and
reduced the amount of fiber recovered after fermentation. Germ
recovery also surprisingly reduced fiber washing and dewatering
difficulties as well.
[0067] Surprisingly protein fraction yields were not significantly
affected by the addition of germ recovery and were found to be 4.0%
of the starting corn. The addition of a fiber wash using the
clarified stillage after centrifugation and protein recovery
surprisingly increased the final protein fraction by 25% to reach a
total yield of 5.1%. This wash could easily be incorporated into a
continuous process without the need for any additional water
usage.
[0068] Compositional Analysis of Coproducts:
[0069] Germ fractions, DDGS equivalents, and protein fractions were
analyzed for protein and lipid content and compared in FIG. 5A and
FIG. 5B as the percentage of the total recovered. The protein
distribution (FIG. 5A) showed surprisingly that the majority of the
protein remained with the DDGS equivalent fractions. The lowest
amount of protein for the DDGS equivalent fractions was found when
both the germ and protein fraction recovery was done and was
reduced further with the addition of the washing step. The wash
removed additional high protein yeast cells as well as insoluble
protein from the fiber. The protein concentration for the recovered
protein fractions ranged from 56.3 to 60.8% on a dry wt basis and
was not found to be significantly different for any of the
treatments. It is likely that the protein concentration may be
altered if an alternative enzyme were used for germ recovery.
[0070] The lipid distribution (FIG. 5B) showed that the
degermination process recovered approximately 65% of the total
lipid in the germ fraction. This value was somewhat lower than
expected given that published values indicate that 80-85% of the
lipid is within the germ (Watson 2003). It is possible that the
difference was due to solvent type or extraction efficiency for the
different lipid components in the germ relative to the endosperm.
It was also possible that some of the germ was not recovered in the
floatation step or that germ was damaged in the grinding operation
and oil leaked into the aqueous medium.
[0071] The DDGS equivalent fractions were also analyzed for fiber
and other feed components and the averaged values are shown in
Table 1 for the five conditions evaluated. The protein
concentration of the DDGS equivalents was surprisingly increased
relative to the conventional controls when germ was recovered;
however, the oil content was reduced significantly. The degermed
DDGS equivalents were reduced in oil concentration by about 50%
relative to the controls. Surprisingly, except for the lipid
content, the overall composition of the DDGS equivalent fractions
were very similar to the conventional DDGS equivalents.
[0072] Overall Economic Evaluation:
[0073] In order to estimate the economic value of our process,
coproduct values and yields were compared between a 100 million
gallon per year conventional facility and a facility producing our
modified coproducts. The comparison of yields and values are shown
in Table 2. The amount of corn utilized for the germ recovery
process was greater relative to the conventional process due to the
slightly lower ethanol yield and resulted in higher overall corn
costs of $3.7 million. The overall value of coproducts was,
however, surprisingly found to overcome the additional corn cost
resulting in almost $17 million in additional revenue. Coproduct
values were based on Midwest pricing given by the USDA Agricultural
Market News for August 2015. The conventional DDGS value was
$140/ton and our DDGS equivalent was estimated to be $95/ton or
slightly lower due to reduced protein and lipid concentrations.
Germ was estimated to be about $300/ton based on oil content and
extraction costs as well as residual extracted meal value. The high
protein fraction was estimated to be $450/ton and was based on the
values of corn gluten meal ($550/ton) and soybean meal
($350/ton).
[0074] Detailed estimates of added operating expenses were not
possible without the development of a full process model; however,
we anticipate that these will surprisingly be less than 5 per
gallon. Without the addition of added operating expenses, the
additional revenue is surprisingly almost 16.7 per gallon of
ethanol produced or about 45 per bushel processed; since the plants
are large, this would be worth about $6 million per year at a 40
million gallon per year plant.
[0075] Conclusions:
[0076] An enzyme based germ and fiber recovery process utilizing a
modified milling strategy has been developed and demonstrated in a
corn ethanol production process. The use of a protease was
demonstrated to yield germ recoveries equivalent to conventional
wet milling. Alpha amylase was also demonstrated to work for germ
recovery; however, other enzymes should also work in this process
if they can alter the specific gravity sufficiently. The ethanol
yield decrease due to fractionation was surprisingly found to be
less than 2.8% relative to conventional controls and could be
reduced further using a continuous process. The DDGS equivalents
produced have significantly reduced lipid content but slightly
higher protein content. Reduced capital input and the high
potential coproduct value should significantly improve the economic
viability for this process relative to previous wet fractionation
methods. Additionally, we have demonstrated that a variation of the
milling process can be used to produce a sugar solution that can
then be utilized in other processes, and another variation would
allow for starch granules to be separated and recovered in a wet
milling type operation.
[0077] Example for Sugar Solution:
[0078] The degerminator ground corn was mixed with water in a 2 L
beaker and the pH adjusted to 4.5 using 1 N HCl. PROSTEEP.RTM. was
added at 0.67 g/kg of mash and the total slurry weight adjusted to
give a solids concentration of 25% or 30% solids. The beaker was
then incubated in a water bath at 48.degree. C. and mixed with a
mechanical mixer for 2 hours. Following incubation, the beaker was
removed and the floating germ recovered by skimming using a small
section of 2 mm wire screen (#10 mesh). The slurry was periodically
mixed to allow more germ to float and again skimmed. This was done
repeatedly until no more germ could be recovered (about 20 min).
The germ was rinsed with 100 mL of water and the rinse water added
to the degermed slurry. The germ collected was dried at 55.degree.
C. in a forced air oven. The degermed corn slurry was readjusted to
25% or 30% solids with water and processed without adjusting the pH
(now about pH 4.9). The particle size of the slurry was further
reduced using an IKA (Wilmington, N.C.) T25 Disperser with an 18G
dispersing element at 16,000 rpm for 3-5 min until the slurry was
uniform. A mechanical mixer was used and Alpha amylase (SPEZYME
RSL.RTM.) was added at 0.5 mL/kg mash and a glucoamylase (OPTIDEX L
400) was added at 0.25 mL/kg mash the slurry incubated at
60.degree. C. for 36 hours. Following incubation, the fiber could
be recovered by screening and the residual solids (primarily
insoluble protein) recovered by centrifugation. The remaining
solution is the sugar solution. Table 3 shows the sugar
concentrations (glucose, maltose, maltotriose and oligosaccharides
of 4 glucose units or larger, (DP4+)) for the 25 and 30% solids
preparations. In each case, the sugar concentrations produced
indicates high yield.
[0079] This example utilized a starch hydrolysis procedure that
does not gelatinize the starch first; however, that process could
also be utilized effectively. This could be accomplished by
incorporating a thermal treatment for gelatinization of the starch
after addition of the alpha amylase but prior to addition of the
glucoamylase. The glucoamylase would then be added after returning
the slurry to 60.degree. C. for incubation. The two processes are
outlined generally in FIG. 2C and FIG. 2D.
[0080] All of the references cited herein, including U.S. patents,
are incorporated by reference in their entirety. Also incorporated
by reference in their entirety are the following references: U.S.
Pat. No. 7,829,680 (Sander, R. and L. D. Fritz (2010); U.S. Pat.
No. 5,231,017 (Lantero, O. J. and J. J. Fish (1993)); Lopes, J. F.,
et al., Cereal Chemistry, 74(5): 633-638 (1997); AACC
International, Approved Methods of Analysis, 11th Ed. (2000), AACC
International, St. Paul, Minn., Method 76-13.01 and 76.11; AOAC,
Official Methods of Analysis of AOAC International (2012), 19th
Ed., AOAC International, Gaithersburg, Md., Methods 930.15, 990.03,
945.16, 942.05, and 985.01; AOCS, Official Methods and Recommended
Practices of the AOCS, 5th ed. (1998), American Oil Chemists'
Society Press, Champaign, Ill., Approved Procedure Ba 6a-05; Wang,
P., et al., Cereal Chemistry, 82(6): 734-738 (2005).
[0081] Thus, in view of the above, there is described (in part) the
following:
[0082] A process of treating grain, said process comprising (or
consisting essentially of or consisting of) milling said grain to
produce milled grain wherein the grain germ remains intact in said
milled grain, and producing a mixture by mixing said milled grain
with water and at least one enzyme selected from the group
consisting of protease, alpha amylase, glucoamylase, cell wall
degrading enzyme, and mixtures thereof, wherein the pH of said
mixture is optionally adjusted to a pH of about 3.5 to about 6.5
(e.g., 3.5 to 6.5), and incubating said mixture for about 1 to
about 3 hours (e.g., 1 to 3 hours; preferably about 2 hours (2
hours)) to produce an incubated mixture (first part of FIG.
2C).
[0083] The above process, further comprising (or consisting
essentially of or consisting of) separating said incubated mixture
into germ and a liquid slurry, reducing the particle size of
materials in said liquid slurry by grinding, adding yeast and
amylase to said liquid slurry and fermenting for about 30 to about
72 hours (e.g., 30 to 72 hours), removing ethanol from said liquid
slurry (e.g., by distillation) and separating (e.g., screening)
said liquid slurry into fiber and a defibered liquid slurry, and
separating (e.g., centrifuging) said defibered liquid slurry to
produce protein solids (balance of FIG. 2A).
[0084] The above process, further comprising (or consisting
essentially of or consisting of) separating said incubated mixture
into germ and a liquid slurry, reducing the particle size of
materials in said liquid slurry by grinding, adding amylase to said
liquid slurry to hydrolyze the starches in said liquid slurry, and
separating (e.g., screening) said liquid slurry into fiber and a
defibered liquid slurry, and separating (e.g., centrifuging]) said
defibered liquid slurry into protein solids and a solution
containing sugars (FIG. 2C).
[0085] The above process, further comprising (or consisting
essentially of or consisting of) separating said incubated mixture
into germ and a liquid slurry, reducing the particle size of
materials in said liquid slurry by grinding, adding amylase to said
liquid slurry and heating (e.g. about 60 to about 130.degree. C.
(60-130.degree. C.), preferably about 75 to about 105.degree. C.
(75-105.degree. C.)) for about 15 min to about 2 hours (e.g., 15
min to 2 hours) (liquefaction), adding glucoamylase enzyme to said
liquid slurry to hydrolyze the starches in said liquid slurry
(saccharification), and separating (e.g., screening) said liquid
slurry into fiber and a defibered liquid slurry, and separating
(e.g., centrifuging) said defibered liquid slurry into protein
solids and a solution containing sugars (FIG. 2D).
[0086] The above process, wherein said grain is corn.
[0087] The term "consisting essentially of" excludes additional
method (or process) steps or composition components that
substantially interfere with the intended activity of the method
(or process) or composition, and can be readily determined by those
skilled in the art (for example, from a consideration of this
specification or practice of the invention disclosed herein).
[0088] The invention illustratively disclosed herein suitably may
be practiced in the absence of any element (e.g., method (or
process) steps or composition components) which is not specifically
disclosed herein.
[0089] Other embodiments of the invention will be apparent to those
skilled in the art from a consideration of this specification or
practice of the invention disclosed herein. It is intended that the
specification and examples be considered as exemplary only, with
the true scope and spirit of the invention being indicated by the
following claims.
TABLE-US-00001 TABLE 1 Average composition of fiber and DDGS
fractions with and without protein fraction recovery.sup.a
Conventional De-Germinated Component Unit With Without With Without
With + Wash Protein (crude) % 27.8 .+-. 1.70 31.6 .+-. 1.41 31.4
.+-. 2.8 36.0 .+-. 2.12 25.5 .+-. 0.78 Fat (crude) % 14.4 .+-. 0.21
13.4 .+-. 1.06 7.43 .+-. 0.8 5.92 .+-. 1.0 4.1 .+-. 0.25 Fiber
(acid detergent) % 9.3 .+-. 0.28 8.6 .+-. 1.70 10.9 .+-. 0.6 10.9
.+-. 0.14 11.0 .+-. 0.21 Fiber (crude) % 5.9 .+-. 0.29 6.0 .+-.
0.42 7.0 .+-. 1.5 6.2 .+-. 0.03 8.6 .+-. 0.36 Fiber (neutral
detergent) % 32.0 .+-. 0.57 27.8 .+-. 0.21 38.0 .+-. 2.3 31.1 .+-.
1.27 35.2 .+-. 0.64 Ash % 5.7 .+-. 0.18 5.4 .+-. 0.22 5.8 .+-. 0.2
5.6 .+-. 0.18 5.9 .+-. 0.04 Hunter color L value L Value 76.3 .+-.
1.6 78.4 .+-. 1.3 76.4 .+-. 1.1 80.4 .+-. 0.21 75.5 .+-. 0.85 Total
digestible nutrients Mcal/lbs 92.9 .+-. 0.6 91.7 .+-. 1.7 84.1 .+-.
0.14 81.8 .+-. 0.57 79.5 .+-. 0.28 Net energy (lactation) Mcal/lbs
1.0 .+-. 0.01 1.0 .+-. 0.02 0.88 .+-. 0.00 0.86 .+-. 0.01 0.8 .+-.
0.00 Net energy (maint.) Mcal/lbs 1.0 .+-. 0.01 1.0 .+-. 0.02 0.92
.+-. 0.00 0.89 .+-. 0.01 0.9 .+-. 0.01 Net energy (gain) Mcal/lbs
0.7 .+-. 0.00 0.7 .+-. 0.01 0.61 .+-. 0.00 0.60 .+-. 0.01 0.6 .+-.
0.00 Digestible energy Mcal/lbs 1.9 .+-. 0.01 1.8 .+-. 0.04 1.68
.+-. 0.00 1.64 .+-. 0.01 1.6 .+-. 0.00 Metabolizable energy
Mcal/lbs 1.7 .+-. 0.01 1.6 .+-. 0.03 1.51 .+-. 0.01 1.45 .+-. 0.00
1.4 .+-. 0.01 Sulfur (total) % 0.3 .+-. 0.01 0.3 .+-. 0.01 0.34
.+-. 0.01 0.39 .+-. 0.02 0.3 .+-. 0.01 Phosphorus (total) % 1.2
.+-. 0.02 1.1 .+-. 0.02 1.08 .+-. 0.06 1.04 .+-. 0.06 0.9 .+-. 0.04
Potassium (total) % 1.4 .+-. 0.00 1.3 .+-. 0.01 1.69 .+-. 0.06 1.53
.+-. 0.04 1.7 .+-. 0.06 Magnesium (total) % 0.4 .+-. 0.01 0.4 .+-.
0.01 0.32 .+-. 0.01 0.3 .+-. 0.01 0.3 .+-. 0.01 Calcium (total) %
0.0 .+-. 0.01 0.0 .+-. 0.01 0.01 .+-. 0.00 0.01 .+-. 0.00 0.04 .+-.
0.01 Sodium (total) % 0.0 .+-. 0.01 0.0 .+-. 0.00 0.06 .+-. 0.00
0.06 .+-. 0.00 0.1 .+-. 0.01 Iron (total) ppm 58.7 .+-. 1.34 71.4
.+-. 0.64 40.4 .+-. 2.40 50.1 .+-. 3.32 35.2 .+-. 2.62 Manganese
(total) ppm 17.8 .+-. 0.28 17.0 .+-. 0.42 12.3 .+-. 0.28 11.6 .+-.
0.57 10.5 .+-. 0.35 Copper (total) ppm 23.7 .+-. 4.60 10.1 .+-.
3.04 31.7 .+-. 11.24 8.9 .+-. 0.71 39.5 .+-. 1.13 Zinc (total) ppm
68.6 .+-. 1.63 65.2 .+-. 3.11 46.9 .+-. 5.73 40.1 .+-. 1.34 44.5
.+-. 0.64 Starch (total) % 2.4 .+-. 0.42 2.5 .+-. 0.04 2.9 .+-.
0.14 2.4 .+-. 0.05 2.6 .+-. 0.02 .sup.aAnalysis was partially done
at a commercial lab for these fractions.
TABLE-US-00002 TABLE 2 Estimated Value of Coproducts for 100
Million Gallon Ethanol Facility.sup.a Conventional De-Germinated
Difference Component Yield Lbs. Value ($) Yield Lbs. Value ($) Lbs.
Value ($) Corn 2,000,000,000 $128,571,429 2,057,613,169
$132,275,132 57,613,169 $3,703,704 DDGS 29.20% 551,555,556
$38,608,889 17.12% 332,693,187 $14,971,193 -218,862,369
-$23,637,695 Germ 0.00% 0.0 0 8.01% 145,173,671 $21,776,051
145,173,671 $21,766,051 Protein 0.00% 0.0 0 5.10% 99,108,368
$22,299,383 99,108,368 $22,299,383 Ethanol 2.80.sup.b 100,000,000
$150,000,000 2.72.sup.b 100,000,000 $150,000,000 0 $0 Output - Corn
$60,037,460 $76,771,495 $16,734,034 cost .sup.aEstimates based on
Corn ($3.6/bushel), DDGS ($140/ton), Ethanol ($1.50/gallon).
Protein price used ($450/ton) was between Soybean meal ($350/ton)
and Corn Gluten Meal ($550/ton). Corn Germ was based on crude oil
price of ($0.41/pound) and germ oil content plus germ meal value.
.sup.bEthanol yields per bushel for Conventional was 2.8 and 2.722
for De-Germinated.
TABLE-US-00003 TABLE 3 Solids Content Sugar 30% 25% DP4.sup.+ 0.40
0.53 Maltotriose 0.51 0.55 Maltose 2.06 2.15 Glucose 24.25
18.73
* * * * *